As disclosed in U.S. Pat. Nos. 5,038,039 and 5,168,162, which are incorporated herein by reference, infrared spectroscopy may be used to detect anomalies in samples of biological tissues and cells in natural and cultured form. A beam of infrared light is directed at the sample, and any anomalies in the sample are detected by analyzing the changes in the infrared absorption within one or more frequency ranges. The change in absorption may evidence itself as a change in absorption intensity at one or more particular frequency absorption peaks, a change in frequency at which one or more absorption peaks occur, the appearance of one or more new absorption peaks at new frequencies, or a combination of two or three of these phenomena. The infrared spectroscopy may be performed at atmospheric pressure or at an elevated pressure. The sample may comprise cells, secretions, exudates, transudates, bacteria, scrapings, brushings or other kinds of exfoliated cells from various organs, tissues or growths including tissue slices, mashed cells, or cells dispersed or suspended in water or other liquids. By using these infrared spectroscopy methods, anomalies such as malignancies and certain diseases, such as diabetes, cirrhosis and arthritis, can be reliably detected at an early stage in their development by persons with little or no medical training. In fact, because the disclosed methods provide numerical results in the form of intensity as a function of frequency, no significant subjective interpretation is required. Consequently, the analysis of samples may be entirely automated and/or computerized, thereby primarily requiring human involvement only in the initial obtaining and preparation of samples for infrared spectral analysis.
Infrared analysis of biological samples is also disclosed in the articles authored by Benedetti et al. in Leukemia Research, Vol. 8, No. 3, pp. 483-489 (1984) and in Leukemia Research, Vol. 9, No. 8, pp. 1001-1008 (1985).
In order to conduct such infrared spectroscopy of such samples, it is typically necessary to mount the sample in some kind of sample holder which has an optical window substantially transparent to the infrared light to be used, which ensures that the infrared light is not completely absorbed by the sample, and which substantially eliminates infrared interference fringes created by the sample which might obscure the sample's infrared absorption spectrum.
An sample holder that may be used at atmospheric pressure in conjunction with the infrared spectroscopy methods described above is disclosed in U.S. Pat. No. 4,980,551, which is incorporated herein by reference. The disclosed sample holder comprises a frame having a passage extending therethrough, and a pair of juxtaposed infrared transparent optical windows disposed within the passage. At least one of the optical windows has a recess formed in the surface abutting the other optical window into which a sample is placed. The sample holder also includes a mask that restricts the infrared light to the recess formed in the optical windows and a means to urge the two optical windows into contact with one another, thereby also clamping the sample within the recess(es). The shape of the recess(es) in the optical window(s) provides paths of propagation of adjacent infrared light rays that are different in length, thereby substantially eliminating interference fringes in the infrared spectra of the sample and clarifying the infrared absorption spectra of the sample being analyzed. The sample holder is useful for examining organ tissue or other deformable semi-solid material, such as, for example, gels, amorphous polymers and highly viscous liquid. Although this sample holder is reliable, functional and useful for many applications, it has certain disadvantages. In particular, after a sample is placed in the recess between the two optical windows, the sample holder must be assembled by mounting and securing the two optical windows into the frame. This assembly process is time consuming and difficult, thereby detracting from the benefits of the infrared spectroscopic analytical method. Additionally, the size of the optical windows in the sample holder is larger than absolutely necessary. Because the infrared absorption spectra is obtained based upon the light that travels through the sample and the optical windows in line therewith, i.e., the portions of the optical windows above and below the recess(es) in the optical window(s), the remaining portions of the optical windows are not needed, as evidenced by the fact that those excess portions of the optical windows are masked. The optical windows are typically the most expensive elements of the sample holder because the material comprising the optical windows is typically expensive and because the optical windows are typically difficult and therefore expensive to fabricate. As a consequence, the inclusion of this excess portion of optical window increases the cost of the sample holder making it unsuitable as a single use, disposable sample holder. The excess portions of the optical windows also make the sample holder less suitable for use in applications where a pressure is to be applied to the sample since a pressure might fracture the optical windows. Additionally, the sample holder typically must be cleaned for reuse, thereby creating a contamination risk, both to the technical personnel involved in the cleaning procedures and to the work environment, and also generating an expensive and logistically difficult problem associated with the safe disposal of the contaminated cleaning agents used in the cleaning procedures.
A sample holder that may be used at high-pressures in conjunction with the above described infrared spectroscopy methods is disclosed in U.S. Pat. No. 4,970,396, which is incorporated herein by reference. This sample holder comprises a frame or gasket made of a high compressive strength material having a passage passing therethrough, and an infrared transparent optical window disposed in the passage. The optical window is sized and shaped so as to form a recess between the upper surface of the optical window and the upper surface of the frame or gasket. A sample can be placed into this recess. As was the case with the above described atmospheric pressure sample holder, the shape of the recess in this sample holder provides paths of propagation of adjacent infrared rays that are different in length to substantially eliminate infrared interference fringes to clarify the infrared absorption spectra. Pressure is applied to the bottom surface of the window and to the sample by anvils which are preferably comprised of diamond crystals. The sample is sealed into the recess of the sample holder when the diamond anvils are pressed into contact with the frame or gasket. This high pressure sample holder is useful for examining liquids, solids, aqueous systems and aqueous biological systems and lipids, proteins, nucleic acids, hydrocarbons and animal or vegetable tissue (alive or dead) and bacteria. Although this sample holder is also reliable, functional and useful for many applications, it also has certain disadvantages. In particular, the anvils are extremely expensive, because the diamonds of which they are comprised are inherently expensive. Fabrication and assembly costs of the sample holder are high because the anvils must be cut and precisely oriented in the sample holder so that the anvil surfaces adjacent to the optical window are parallel to one another within a tolerance of one interference fringe for the particular frequency of light being used in the interferometer. Additionally, because the optical path of the diamond anvils is relatively small (less than about 0.5 mm.), in order to obtain reasonably acceptable infrared spectra, it is highly desirable to use a light beam condenser and a highly sensitive infrared detector. As a consequence of using these supplementary elements, proper optical alignment among the elements and the sample is difficult to obtain. The disclosed sample holder also cannot be used to obtain infrared spectra at atmospheric pressure. Finally, because of the relatively high cost associated with manufacturing this particular sample holder, it is not suitable as a single use, disposable sample holder, but must be cleaned for reuse thereby creating the contamination and disposal problems discussed above.